Distribution of the 5-hydroxytryptamine2C receptor protein in adult rat brain and spinal cord determined using a receptor-directed antibody: Effect of 5,7-dihydroxytryptamine
✍ Scribed by Anita Sharma; Taniya Punhani; Kevin C.F. Fone
- Publisher
- John Wiley and Sons
- Year
- 1997
- Tongue
- English
- Weight
- 296 KB
- Volume
- 27
- Category
- Article
- ISSN
- 0887-4476
No coin nor oath required. For personal study only.
✦ Synopsis
A synthetic peptide, corresponding to the N-terminal decapeptide (1Y 11 C 12 ) of the rat 5-hydroxytryptamine 2C (5-HT 2C ) receptor protein was used to produce a sheep polyclonal antiserum. Western blot analysis showed that the resultant antibody G241 recognised two membrane proteins, one (55 kDa) approximating the molecular mass of the 5-HT 2C receptor (52 kDa) and a second (63 kDa), which may be a glycosylated form of the receptor protein. HEK 293 cells transfected with human 5-HT 2C cDNA displayed intense cell surface immunoreactivity with the 5-HT 2C antiserum, which was completely prevented by incubating the antibody with the synthetic 5-HT 2C peptide (10 µM), whilst neither nonimmune serum nor untransfected cells displayed any immunoreactivity. A radioimmunoassay was developed to quantify the regional distribution of 5-HT 2C -like immunoreactivity (LI) in the adult rat brain. The choroid plexus contained five-fold higher levels of 5-HT 2C -LI than any brain region but high levels were found in the frontal cortex, septum, hypothalamus, and striatum, intermediate levels in the thalamus and midbrain, and lower levels in brainstem, cerebellum, and spinal cord. In rat cortical membranes, the B max value from [ 3 H]-mesulergine binding was ten-fold lower than 5-HT 2C -LI levels determined by radioimmunoassay, which may reflect measurement of internalised receptor protein by radioimmunoassay which is not detected with conventional 5-HT 2C ligands. Ten days after depletion of 5-HT with the serotonergic neurotoxin 5,7dihydroxytryptamine (5,7-DHT), there was a significant increase in 5-HT 2C -LI in the choroid plexus and the ventral cervical spinal cord, suggesting that receptors therein are located post-synaptic to destroyed serotonergic nerve terminals. In contrast, the significant reduction in 5-HT 2C -LI observed in the midbrain, brainstem, and dorsal thoracic spinal cord following 5,7-DHT implies that 5-HT 2C receptors may be located on 5-HT nerve terminals in these regions.